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A novel mammalian protein that may promote apoptosis by binding to and antagonizing IAP proteins has been identified. It is called DIABLO for 'direct IAP-binding protein with low pl.' Coimmunoprecipitation and 2D immobilized pH gradient/SDS PAGE was used, followed by electrospray ionization tandem mass spectrometry.

Aflatoxin B1 (AFB1), ochratoxin A (OTA) and fumonisin B1 (FB1) are contaminants which have been shown to regularly co-occur in a range of foods. However, only a small number of studies have evaluated the interactive effect of binary and tertiary mycotoxins. The present study evaluated the effects of low levels of each mycotoxin in combination at their EU regulatory limits. Toxic effect with respect to cell viability was measured by MTT and neutral red assays, assessing mitochondria and lysosome integrities respectively. Individual toxicity showed that OTA (10 μg/ml) was the most cytotoxic mycotoxin in all three cell lines studied (caco-2, MDBK and raw 264.7). Binary combinations were cytotoxic to the MDBK cell line in the order [OTA/FB1] > [AFB1/FB1] > [AFB1/OTA], whilst all effects observed were classified as being additive. Tertiary combinations of AFB1, FB1 and OTA at the EU regulatory limits were tested and not found to exhibit measurable cytotoxicity in MDBK, caco-2 or raw 264.7 cells. However by increasing these concentrations above the legal limits to OTA (3 μg/ml), FB1 (8 μg/ml) and AFB1 (1.28 μg/ml), cytotoxicity was observed with up to 26% reduction in cell viability and synergistic effects were evident with regard to mitochondrial integrity. [ABSTRACT FROM AUTHOR]

• The insulin-like growth factor system can be impacted by endocrine disruptors. • Prenatal disruption of insulin-like growth factor can affect fetal growth. • Dioxins and dioxin-like compounds affect the system in experimental models. • Phthalates adversely affect the system in human studies. The insulin-like growth factor (IGF) system is a critical regulator of growth, especially during fetal development, while also playing a central role in metabolic homeostasis. Endocrine disruptors (EDs) are ubiquitous compounds able to interfere with hormone action and impact human health. For example, exposure to EDs is associated with decreased birthweight and increased incidence of metabolic disorders. Therefore, the IGF system is a potential target for endocrine disruption. This review summarises the state of the science regarding effects of exposure to major classes of endocrine disruptors (dioxins and dioxin-like compounds, polycyclic aromatic hydrocarbons, polybrominated diphenyl ethers, phthalates, perfluoroalkyl substances and bisphenol A) on the IGF system. Evidence from both experimental models (in vitro and in vivo) and epidemiological studies is presented. In addition, possible molecular mechanisms of action and effects on methylation are discussed. There is a large body of evidence supporting the link between dioxins and dioxin-like compounds and IGF disruption, but mixed findings have been reported in human studies. On the other hand, although only a few animal studies have investigated the effects of phthalates on the IGF system, their negative association with IGF levels and methylation status has been more consistently reported in humans. For polybrominated diphenyl ethers, perfluoroalkyl substances and bisphenol A the evidence is still limited. Despite a lack of studies for some ED classes linking ED exposure to changes in IGF levels, and the need for further research to improve reproducibility and determine the degree of risk posed by EDs to the IGF system, this is clearly an area of concern. [ABSTRACT FROM AUTHOR]

Patulin (PAT) is a mycotoxin produced by various species of fungi, with Penicillium expansum being the most commonly occurring. Apples and apple products are the main sources of PAT contamination. This mycotoxin has been shown to induce toxic effects in animals, a few of which include reproductive toxicity and interference with the endocrine system. Here the endocrine disrupting potential of PAT has been investigated in vitro to identify disruption at the level of oestrogen, androgen, progestagen and glucocorticoid nuclear receptor transcriptional activity, and to assess interferences in estradiol, testosterone and progesterone steroid hormone production. At the receptor level, 0.5-5000 ng/ml (0.0032-32 μM) PAT did not appear to induce any specific (ant) agonistic responses in reporter gene assays (RGAs); however, nuclear transcriptional activity was affected. A >6 fold increase in the glucocorticoid receptor transcriptional activity was observed following treatment with 5000 ng/ml PAT in the presence of cortisol. At the hormone production level, despite cytotoxicity being observed after treatment with 5000 ng/ml PAT, estradiol levels had increased >2 fold. At 500 ng/ml PAT treatment, an increase in progesterone and a decrease in testosterone production were observed. The findings of this study could be considered in assessing the health risks following exposure to PAT. [ABSTRACT FROM AUTHOR]

• Nerve growth factor (NGF) induced neuritogenesis in PC12 rat pheochromocytoma cells. • Persistent organic pollutants (POPs) enhanced NGF-induced neuritogenesis. • Perfluorooctane sulfonic acid (PFOS) contributed to 50 % of the POP mixture effect. • Nuclear and mitochondrial health was unaffected by the POP mixture or PFOS. Disruption of neurite outgrowth is a marker for neurotoxicity. Persistent organic pollutants (POPs) are potential developmental neurotoxicants. We investigated their effect on neurite outgrowth in PC12 rat pheochromocytoma cells, in absence or presence of nerve growth factor (NGF), an inducer of neuronal differentiation. Cells were exposed for 72 h to a defined mixture of POPs with chemical composition and concentrations based on blood levels in the Scandinavian population. We also evaluated perfluorooctane sulfonic acid (PFOS) alone, the most abundant compound in the POP mixture. Only higher concentrations of POP mixture reduced tetrazolium salt (MTT) conversion. High-content analysis showed a decrease in cell number, but no changes for nuclear and mitochondrial cellular health parameters. Robust glutathione levels were observed in NGF-differentiated cells. Live imaging, using the IncuCyte ZOOM platform indicated ongoing cell proliferation over time, but slower in presence of NGF. The pollutants did not inhibit neuritogenesis, but rather increased NGF-induced neurite length. PFOS induced neurite outgrowth to about 50 % of the level seen with the POP mixture. Neither the POP mixture nor PFOS affected neurite length in the absence of NGF. Our observations indicate that realistic complex mixtures of environmental pollutants can affect neuronal connectivity via NGF-induced neurite outgrowth. [ABSTRACT FROM AUTHOR]

The mycotoxins zearalenone (ZEN) and alpha-zearalenone (α-ZOL), which are common contaminants of agri-food products, are known for their oestrogenic potential. In addition to mycotoxins, food may also contain pesticides with oestrogenic properties such as 1,1,1-trichloro-2,2-bis(p-chlorophenyl) ethane (p,p' -DDT) and 1,1-dichloro-2,2-bis(p -chlorophenyl) ethylene (p,p' -DDE), raising the question on the potential effects of individual and combinations of these xeno-oestrogens on the action of natural oestrogens. The present study employed a mammalian reporter gene assay to assess the effects individual and binary combinations of these environmental and food-borne contaminants on oestrogen nuclear receptor (ER) transactivation. As expected, α-ZOL and ZEN exhibited the strongest oestrogenic potency (EC 50 : 0.27 ± 0.121 nM and 1.32 ± 0.0956 nM, respectively) whereas p,p' -DDT and p,p' -DDE had weak ER agonistic activity with the maximal response of 28.70 ± 2.97% and 18.65 ± 1.77%, respectively. Concurrent treatment of the mycotoxins and/or pesticides, individually or in binary combination, with 17β-oestradiol (E 2) showed either additive, synergistic or antagonistic interactive effects on E 2 -mediated ER response, depending on the combination ratios, the concentration range of xeno-oestrogens, and the concentration of E 2. This study highlights the importance of assessing the mixture effects of chemical contaminants in risk assessment, especially in the area of reproductive and developmental toxicity. • The effects of individual and binary mixtures of mycotoxins and pesticides on oestrogen receptor (ER) transcription response was evaluated. • Zearalenone (ZEN) and α-zearalenol (α-ZOL) exhibited the strong oestrogenic potency. • ZEN or α-ZOL, alone or in combination, antagonised the ER response mediated by 10 nM of 17β-oestradiol (E 2). • Binary combination of ZEN or α-ZOL with either 1,1,1-trichloro-2,2-bis(p -chlorophenyl) ethane (p,p' -DDT) and 1,1-dichloro-2,2-bis(p -chlorophenyl) ethylene (p,p' -DDE) antagonised the ER response mediated by E 2. • Equimolar concentrations of p,p' -DDT and p,p' -DDE at 50 μM completely inhibited the ER response mediated by E 2. [ABSTRACT FROM AUTHOR]

Abstract Global market of herbs has been struggling with food adulteration issues. A number of assays have been developed to aid the detection of the tampered samples and ensure high quality of the marketed products. However, herbs are marketed not only for their culinary applications but also as remedies due to high levels of biologically active constituents. Nevertheless, there is no information in the literature about the influence of herbs adulteration on the biological activity of the final product. Current study aims at assessing the influence of oregano adulteration on its in-vitro estrogen-like activity. High responses in a mammalian reporter gene assay have been detected in pure and adulterated samples, translating to 21–7409 ng of 17β-estradiol equivalents per gram of oregano. The origin of those responses was assessed by combining fractionation and UHPLC-HRMS. Three flavones were proposed as the most active extract constituents i.e. luteolin-glucoside, luteolin- and apigenin-glucuronides all of which have been previously identified in other herbal extracts with estrogenic activity. This study underlines challenges of biological activity assessment in complex herbal extracts as well as the need for further assessment of such supplement administrations in the case of postmenopausal women and breast cancer patients undergoing hormone therapy. Highlights • Oregano extracts exhibit estrogen-like activity in-vitro. • Conjugated flavonoids are the origin of the detected biological activity. • Combining bio- and chemical assays is a fast tool for detecting bioactive compounds. [ABSTRACT FROM AUTHOR]

Highlights • Monosodium glutamate (MSG) is a suspected obesogen. • There is a link between exposure to MSG and alterations in steroid hormone levels. • MSG can antagonise the androgen receptor in a dose dependent manner. • The androgen receptor may be an emerging target for diabetes and obesity. Abstract Monosodium glutamate (MSG) is a commonly used flavour enhancer in households, catering and food production. Recently it has been highlighted as a suspected dietary obesogen in epidemiological studies indicating a link between MSG consumption and weight gain. Additionally, animal studies have shown that MSG exposure has profound effects on sex steroid hormone levels and receptors; which have an important role in energy metabolism. However, the exact mechanism by which MSG exerts its effects has yet to be elucidated. Reporter gene assays (RGAs) and the H295R steroidogenesis assay have been used to investigate the endocrine disrupting potential of MSG. Receptor (ant)agonism was not observed in the MMV-Luc (oestrogen responsive) or TM-Luc (progestagen responsive) cell lines following exposure to MSG. Also, no effects on hormone production were observed. However, MSG exhibited an antagonist response in the androgen and progestagen responsive TARM-Luc cell line, with a dose dependent reduction in androgen response of 33%, 36.9% and 50.6% (in comparison to the solvent control) at 50, 250 and 500 μg/ml MSG, respectively (P ≤ 0.05; P ≤ 0.05; P ≤ 0.001). No cytotoxicity or pre-lethal cytotoxicity was observed at the concentrations tested. These findings demonstrate one potential pathway whereby MSG may act as a dietary obesogen. [ABSTRACT FROM AUTHOR]

Monosodium glutamate (MSG) is a suspected obesogen with epidemiological evidence positively correlating consumption to increased body mass index and higher prevalence of metabolic syndrome. ELISA and high content analysis (HCA) were employed to examine the disruptive effects of MSG on the secretion of enteroendocrine hormone glucagon-like peptide-1 (GLP-1) and GLP-1 receptor (GLP-1R), respectively. Following 3 h MSG exposure of the enteroendocrine pGIP/neo: STC-1 cell line model (500 μg/ml) significantly increased GLP-1 secretion (1.8 fold; P ≤ 0.001), however, 72 h exposure (500 μg/ml) caused a 1.8 fold decline (P ≤ 0.05). Also, 3 h MSG exposure (0.5–500 μg/ml) did not induce any cytotoxicity (including multiple pre-lethal markers) but 72 h exposure at 250–500 μg/ml, decreased cell number (11.8–26.7%; P ≤ 0.05), increased nuclear area (23.9–29.8%; P ≤ 0.001) and decreased mitochondrial membrane potential (13–21.6%; P ≤ 0.05). At 500 μg/ml, MSG increased mitochondrial mass by 16.3% (P ≤ 0.01). MSG did not agonise or antagonise internalisation of the GLP-1R expressed recombinantly in U2OS cells, following GLP-1 stimulation. In conclusion, 72 h exposure of an enteroendocrine cell line at dietary levels of MSG, results in pre-lethal cytotoxicity and decline in GLP-1 secretion. These adverse events may play a role in the pathogenesis of obesity as outlined in the obesogen hypothesis by impairing GLP-1 secretion, related satiety responses and glucose-stimulated insulin release. [ABSTRACT FROM AUTHOR]

Fumonisin B1 (FB1) and beauvericin (BEA) are secondary metabolites of filamentous fungi, which under appropriate temperature and humidity conditions may develop on various foods and feeds. To date few studies have been performed to evaluate the toxicological and endocrine disrupting effects of FB1 and BEA. The present study makes use of various in vitro bioassays including; oestrogen, androgen, progestagen and glucocorticoid reporter gene assays (RGAs) for the study of nuclear receptor transcriptional activity, the thiazolyl blue tetrazolium bromide (MTT) assay to monitor cytotoxicity and high content analysis (HCA) for the detection of pre-lethal toxicity in the RGA and Caco-2 human colon adenocarcinoma cells. At the receptor level, 0.001–10 μM BEA or FB1 did not induce any agonist responses in the RGAs. However at non-cytotoxic concentrations, an antagonistic effect was exhibited by FB1 on the androgen nuclear receptor transcriptional activity at 10 μM and BEA on the progestagen and glucocorticoid receptors at 1 μM. MTT analysis showed no decrease in cell viability at any concentration of FB1, whereas BEA showed a significant decrease in viability at 10 μM. HCA analysis confirmed that the reduction in the progestagen receptor transcriptional activity at 1 μM BEA was not due to pre-lethal toxicity. In addition, BEA (10 μM) induced significant toxicity in both the TM-Luc (progestagen responsive) and Caco-2 cells. [ABSTRACT FROM AUTHOR]

The food industry is moving towards the use of natural sweeteners such as those produced by Stevia rebaudiana due to the number of health and safety concerns surrounding artificial sweeteners. Despite the fact that these sweeteners are natural; they cannot be assumed safe. Steviol glycosides have a steroidal structure and therefore may have the potential to act as an endocrine disruptor in the body. Reporter gene assays (RGAs), H295R steroidogenesis assay and Ca 2+ fluorimetry based assays using human sperm cells have been used to assess the endocrine disrupting potential of two steviol glycosides: stevioside and rebaudioside A, and their metabolite steviol. A decrease in transcriptional activity of the progestagen receptor was seen following treatment with 25,000 ng/ml steviol in the presence of progesterone (157 ng/ml) resulting in a 31% decrease in progestagen response ( p = <0.01). At the level of steroidogenesis, the metabolite steviol (500–25,000 ng/ml) increased progesterone production significantly by 2.3 fold when exposed to 10,000 ng/ml ( p = <0.05) and 5 fold when exposed to 25,000 ng/ml ( p =<0.001). Additionally, steviol was found to induce an agonistic response on CatSper, a progesterone receptor of sperm, causing a rapid influx of Ca 2+ . The response was fully inhibited using a specific CatSper inhibitor. These findings highlight the potential for steviol to act as a potential endocrine disruptor. [ABSTRACT FROM AUTHOR]

Persistent organic pollutants (POPs) are toxic substances, highly resistant to environmental degradation, which can bio-accumulate and have long-range atmospheric transport potential ( UNEP, 2001 ). The majority of studies on endocrine disruption have focused on interferences on the sexual steroid hormones and so have overlooked disruption to glucocorticoid hormones. Here the endocrine disrupting potential of individual POPs and their mixtures has been investigated in vitro to identify any disruption to glucocorticoid nuclear receptor transcriptional activity. POP mixtures were screened for glucocorticoid receptor (GR) translocation using a GR redistribution assay (RA) on a CellInsight™ NXT high content screening (HCS) platform. A mammalian reporter gene assay (RGA) was then used to assess the individual POPs, and their mixtures, for effects on glucocorticoid nuclear receptor transactivation. POP mixtures did not induce GR translocation in the GR RA or produce an agonist response in the GR RGA. However, in the antagonist test, in the presence of cortisol, an individual POP, p , p ′-dichlorodiphenyldichloroethylene ( p , p′ -DDE), was found to decrease glucocorticoid nuclear receptor transcriptional activity to 72.5% (in comparison to the positive cortisol control). Enhanced nuclear transcriptional activity, in the presence of cortisol, was evident for the two lowest concentrations of perfluorodecanoic acid (PFOS) potassium salt (0.0147 mg/ml and 0.0294 mg/ml), the two highest concentrations of perfluorodecanoic acid (PFDA) (0.0025 mg/ml and 0.005 mg/ml) and the highest concentration of 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47) (0.0000858 mg/ml). It is important to gain a better understanding of how POPs can interact with GRs as the disruption of glucocorticoid action is thought to contribute to complex diseases. [ABSTRACT FROM AUTHOR]

Zearalenone (ZEN) is a mycotoxin produced by Fusarium fungi. Once ingested, ZEN may be absorbed and metabolised to α- and β-zearalenol (α-ZOL, β-ZOL), and to a lesser extent α- and β-zearalanol (α-ZAL, β-ZAL). Further biotransformation to glucuronide conjugates also occurs to facilitate the elimination of these toxins from the body. Unlike ZEN and its metabolites, information regarding the estrogenic activity of these glucuronide conjugates in various tissues is lacking. ZEN-14- O -glucuronide, α-ZOL-14- O -glucuronide, α-ZOL-7- O -glucuronide, β-ZOL-14- O -glucuronide and β-ZOL-16- O -glucuronide, previously obtained as the major products from preparative enzymatic synthesis, were investigated for their potential to cause endocrine disruption through interference with estrogen receptor transcriptional activity. All five glucuronide conjugates showed a very weak agonist response in an estrogen responsive reporter gene assay (RGA), with activity ranging from 0.0001% to 0.01% of that of 17β-estradiol, and also less than that of ZEN, α-ZOL and β-ZOL which have previously shown estrogenic potencies of the order 17β-estradiol > α-ZOL > ZEN > β-ZOL. Confirmatory mass spectrometry revealed that any activity observed was likely a result of minor deconjugation of the glucuronide moiety. This study confirms that formation of ZEN and ZOL glucuronides is a detoxification reaction with regard to estrogenicity, serving as a potential host defence mechanism against ZEN-induced estrogenic activity. [ABSTRACT FROM AUTHOR]

Evidence that some of the fungal metabolites present in food and feed may act as potential endocrine disruptors is increasing. Enniatin B (ENN B) is among the emerging Fusarium mycotoxins known to contaminate cereals. In this study, the H295R and neonatal porcine Leydig cell (LC) models, and reporter gene assays (RGAs) have been used to investigate the endocrine disrupting activity of ENN B. Aspects of cell viability, cell cycle distribution, hormone production as well as the expression of key steroidogenic genes were assessed using the H295R cell model. Cell viability and hormone production levels were determined in the LC model, while cell viability and steroid hormone nuclear receptor transcriptional activity were measured using the RGAs. ENN B (0.01–100 μM) was cytotoxic in the H295R and LC models used; following 48 h incubation with 100 μM. Flow cytometry analysis showed that ENN B exposure (0.1–25 μM) led to an increased proportion of cells in the S phase at higher ENN B doses (>10 μM) while cells at G 0 /G 1 phase were reduced. At the receptor level, ENN B (0.00156–15.6 μM) did not appear to induce any specific (ant) agonistic responses in reporter gene assays (RGAs), however cell viability was affected at 15.6 μM. Measurement of hormone levels in H295R cells revealed that the production of progesterone, testosterone and cortisol in exposed cells were reduced, but the level of estradiol was not significantly affected. There was a general reduction of estradiol and testosterone levels in exposed LC. Only the highest dose (100 μM) used had a significant effect, suggesting the observed inhibitory effect is more likely associated with the cytotoxic effect observed at this dose. Gene transcription analysis in H295R cells showed that twelve of the sixteen genes were significantly modulated ( p < 0.05) by ENN B (10 μM) compared to the control. Genes HMGR, StAR, CYP11A, 3βHSD2 and CYP17 were downregulated, whereas the expression of CYP1A1, NR0B1, MC2R, CYP21, CYP11B1, CYP11B2 and CYP19 were upregulated. The reduction of hormones and modulation of genes at the lower dose (10 μM) in the H295R cells suggests that adrenal endocrine toxicity is an important potential hazard. [ABSTRACT FROM AUTHOR]

Highlights: [•] Estrogenic activity is quantified in sports supplements and an exposure and risk assessment performed. [•] 17β-Estradiol equivalent activity levels were higher than the ADI in 26% of exposure assessment samples. [•] 17β-Estradiol equivalent activity levels were higher in 66% of exposure assessment samples than levels present in the typical diet. [•] The predicted hormonal impact of greatest concern is for young boys and postmenopausal women. [•] Consumers of sport supplements may be exposed to high levels of estrogenic endocrine disruptors. [Copyright &y& Elsevier]

Reducing exposure to obesogens is a strategy for preventing obesity. [Display omitted] • There is an expanding global obesity pandemic. • Ubiquitous environmental chemicals called obesogens play a vital role in the obesity pandemic. • Exposure to obesogens occurs throughout the life course from before conception until death. • Development is the most sensitive time for obesogens to impact future weight gain across the lifespan and generations. • Obesogens can act via epigenetic mechanisms. • There is a need to expand understanding of the obesogen paradigm to clinicians and consumers. Obesity is a multifactorial disease with both genetic and environmental components. The prevailing view is that obesity results from an imbalance between energy intake and expenditure caused by overeating and insufficient exercise. We describe another environmental element that can alter the balance between energy intake and energy expenditure: obesogens. Obesogens are a subset of environmental chemicals that act as endocrine disruptors affecting metabolic endpoints. The obesogen hypothesis posits that exposure to endocrine disruptors and other chemicals can alter the development and function of the adipose tissue, liver, pancreas, gastrointestinal tract, and brain, thus changing the set point for control of metabolism. Obesogens can determine how much food is needed to maintain homeostasis and thereby increase the susceptibility to obesity. The most sensitive time for obesogen action is in utero and early childhood, in part via epigenetic programming that can be transmitted to future generations. This review explores the evidence supporting the obesogen hypothesis and highlights knowledge gaps that have prevented widespread acceptance as a contributor to the obesity pandemic. Critically, the obesogen hypothesis changes the narrative from curing obesity to preventing obesity. [ABSTRACT FROM AUTHOR]

Abstract: A panel of reporter gene assays (RGAs) coupled with a single solid phase extraction (SPE) step was developed and used to screen bottled mineral water for the presence of four classes of endocrine disruptors (EDs), oestrogens, androgens, progestagens and glucocorticoids. Fourteen brands of bottled mineral water in triplicate (42 samples) were analysed. Overall, hormonal activity was found in 78% of the samples. Oestrogenic, androgenic, progestagenic and glucocorticoid activity was found in 38%, 38%, 36% and 55% of the samples, respectively at an average concentration of 10ng/l 17β-estradiol equivalent (EEQ), 26ng/l testosterone equivalent (TEQ), 123ng/l progesterone equivalent (PEQ) and 13.5ng/l hydrocortisone equivalent (HEQ). The level of oestrogenic, androgenic and progestagenic activity observed is not considered a matter of concern for the consumers’ health. It is unknown whether the glucocorticoid levels observed are safe. The ED source, long term exposure and mixture effects remain to be investigated. [Copyright &y& Elsevier]

Abstract: Pantothenic acid (PA), vitamin B5, is an essential B vitamin that may be fortified in food and as such requires robust and accurate methods of detection to meet compliance legislation. This study reports the production and characterisation of the first monoclonal antibody (MAb) specific for PA and the subsequent development of a surface plasmon resonance (SPR) biosensor assay for the quantification of PA. The developed assay was compared with an SPR based commercial kit which utilised a polyclonal antibody (PAb). Foodstuffs, including cereals (n =43), infant formulas and baby food (n =10) and fruit juices (n =48) were analysed by both the MAb and PAb biosensor assays and comparison plots showed good correlation (R 2 0.77–0.99). The results indicate that the MAb based biosensor assay is suitable for the measurement of PA in foodstuffs and has the added advantage of facilitating a constant, long term supply of identical antibody. Preliminary matrix studies suggest the MAb based assay is an excellent candidate for further validation studies and routine quality assurance based analysis. [Copyright &y& Elsevier]

Abstract: Constructed wetland systems (CWS) have been used as a low cost bio-filtration system to treat farm wastewater. While studies have shown that CWS are efficient in removing organic compounds and pathogens, there is limited data on the presence of hormones in this type of treatment system. The objective of this study was to evaluate the ability of the CWS to reduce estrogenic and androgenic hormone concentration in dairy wastewater. This was achieved through a year long study on dairy wastewater samples obtained from a surface flow CWS. Analysis of hormonal levels was performed using a solid phase extraction (SPE) sample clean-up method, combined with reporter gene assays (RGAs) which incorporate relevant receptors capable of measuring total estrogenic or androgenic concentrations as low as 0.24 ng L−1 and 6.9 ng L−1 respectively. Monthly analysis showed a mean removal efficiency for estrogens of 95.2%, corresponding to an average residual concentration of 3.2 ng L−1 17β-estradiol equivalent (EEQ), below the proposed lowest observable effect concentration (LOEC) of 10 ng L−1. However, for one month a peak EEQ concentration of 115 ng L−1 was only reduced to 18.8 ng L−1. The mean androgenic activity peaked at 360 ng L−1 and a removal efficiency of 92.1% left an average residual concentration of 32.3 ng L−1 testosterone equivalent (TEQ). The results obtained demonstrate that this type of CWS is an efficient system for the treatment of hormones in dairy wastewater. However, additional design improvements may be required to further enhance removal efficiency of peak hormone concentrations. [Copyright &y& Elsevier]

Abstract: α- and β-zearalenol (α-ZOL and β-ZOL, respectively) are metabolites of the mycotoxin zearalenone (ZEN). All three individual mycotoxins have shown to be biological active i.e. being estrogenic and able to stimulate cellular proliferation albeit at different strengths. In this work, cytosol protein expression was determined by using stable-isotope labelling by amino acids in cell culture (SILAC) upon exposure of α-ZOL and β-ZOL to the steroidogenesis cell model H295R. A total of 14 and 5 individual proteins were found to be significantly regulated by α-ZOL and β-ZOL, respectively. Interestingly, there were no common protein regulations by the metabolites or the parent mycotoxin ZEN. Furthermore, the regulated proteins were assigned to networks and groups of actions that also differed from one another suggesting that the three individual mycotoxins may have unique biological activities. [Copyright &y& Elsevier]

Abstract: Zearalenone (ZEN) is a mycotoxin with endocrine disrupting effects having vast economic implications in e.g. pig farming. Structurally, ZEN resembles 17β-estradiol, and thus is able to bind to estrogen receptors (ER) in target cells. Because of this, it is also classified as a non-steroidal estrogen, a phytoestrogen, a mycoestrogen, and a growth promoter. Quantitative proteomic analysis was undertaken using stable-isotope labeling by amino acids in cell culture (SILAC) upon exposure of the steroidogenesis cell model H295R with ZEN to elucidate its effect on protein regulation. ZEN significantly regulated 21 proteins, including proteins with known endocrine disrupting effects and several oncogenes. In addition, network analysis using Ingenuity Pathway Analysis showed that ZEN affected the oxidative phosphorylation pathway and the mitochondrial dysfunction pathway, both previously reported to be involved in endocrine dysfunction. [Copyright &y& Elsevier]

Abstract: Okadaic acid (OA) and structurally related toxins dinophysistoxin-1 (DTX-1), and DTX-2, are lipophilic marine biotoxins. The current reference method for the analysis of these toxins is the mouse bioassay (MBA). This method is under increasing criticism both from an ethical point of view and because of its limited sensitivity and specificity. Alternative replacement methods must be rapid, robust, cost effective, specific and sensitive. Although published immuno-based detection techniques have good sensitivities, they are restricted in their use because of their inability to: (i) detect all of the OA toxins that contribute to contamination; and (ii) factor in the relative toxicities of each contaminant. Monoclonal antibodies (MAbs) were produced to OA and an automated biosensor screening assay developed and compared with ELISA techniques. The screening assay was designed to increase the probability of identifying a MAb capable of detecting all OA toxins. The result was the generation of a unique MAb which not only cross-reacted with both DTX-1 and DTX-2 but had a cross-reactivity profile in buffer that reflected exactly the intrinsic toxic potency of the OA group of toxins. Preliminary matrix studies reflected these results. This antibody is an excellent candidate for the development of a range of functional immunochemical-based detection assays for this group of toxins. [Copyright &y& Elsevier]

The suppressors of cytokine signaling (SOCS) proteins inhibit cytokine action by direct interaction with Janus kinases or activated cytokine receptors. In addition to the N-terminal and Src homology 2 domains that mediate these interactions, SOCS proteins contain a C-terminal SOCS box. DNA data base searches have identified a number of other protein families that possess a SOCS box, of which the ankyrin repeat and SOCS box-containing (Asb) proteins constitute the largest. Although it is known that the SOCS proteins are involved in the negative regulation of cytokine signaling, the biological and biochemical functions of the Asbs are largely undefined. Using a proteomics approach, we demonstrate that creatine kinase B (CKB) interacts with Asb-9 in a specific, SOCS box-independent manner. This interaction increases the polyubiquitylation of CKB and decreases total CKB levels within the cell. The targeting of CKB for degradation by Asb-9 was primarily SOCS box-dependent and suggests that Asb-9 acts as a specific ubiquitin ligase regulating levels of this evolutionarily conserved enzyme. [ABSTRACT FROM AUTHOR]